Nicotinic cholinergic regulation of tyrosine hydroxylase gene expression and catecholamine synthesis in isolated bovine adrenal chromaffin cells.
Nicotinic cholinergic regulation of tyrosine hydroxylase gene expression and catecholamine synthesis in isolated bovine adrenal chromaffin cells.
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分离牛肾上腺嗜铬细胞中酪氨酸羟化酶基因表达和儿茶酚胺合成的烟碱胆碱能调节。
DOI：
10.1111/j.1471-4159.1992.tb10122.x
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发表时间：
1992
影响因子：
4.7
通讯作者：
Waymire,JC
中科院分区：
文献类型：
--
作者：
Craviso,GL;Hemelt,VB;Waymire,JC
用离体牛肾上腺嗜铬细胞研究烟碱对酪氨酸羟化酶（TH）基因表达的调控。细胞连续暴露于卡巴胆碱或烟碱受体激动剂1，1-二甲基-4-苯基哌嗪（DMPP）可导致TH酶活性呈时间和浓度依赖性增加，而毒蕈碱则无影响。在存在2.5 mM钙的情况下，1 μM（EC 50 = 0.3 μM）的DMPP在3-5天后引起TH活性和TH免疫反应蛋白水平升高两到三倍;在较低的细胞外钙水平下，酶水平的增加显着较小。酪氨酸羟化为多巴胺（DA）在完整的细胞，内源性TH活性的指数，增加与TH水平的上升平行的速率。TH mRNA水平在蛋白水平增加之前升高。如核运行试验所确定，向细胞中添加1 μMDMPP后30分钟内，TH基因转录被刺激两到三倍;转录在2小时内恢复到基础水平。尼群地平（20 μM）阻断DMPP对转录的刺激。用放线菌酮（5 μM）预处理细胞不会阻止DMPP对转录的刺激。毛喉素（10 μM）也通过一种不被放线菌酮阻断的机制增加TH转录（15分钟内增加4倍）。这些结果表明，烟碱受体刺激以钙依赖性方式增加TH mRNA合成、TH蛋白水平和TH活性。此外，烟碱对TH基因表达的影响似乎并不需要蛋白质因子的合成来发挥其作用。原位DA合成率随着TH水平的升高而升高，这表明TH诱导是一种长期增强嗜铬细胞儿茶酚胺合成能力的机制。
Isolated bovine adrenal chromaffin cells were used to study the nicotinic regulation of tyrosine hydroxylase (TH) gene expression. Continuous exposure of the cells to carbachol or the nicotinic receptor agonist 1,1‐dimethyl‐4‐phenylpiperazinium (DMPP) produces a time‐ and concentration‐dependent increase in TH enzyme activity, whereas muscarine has no effect. DMPP at 1 μM(EC50= 0.3 μM) elicits a two‐ to threefold elevation of both TH activity and TH immunoreactive protein level after 3–5 days in the presence of 2.5 mMcalcium; the increase in enzyme levels is significantly less at lower extracellular calcium levels. The rate of hydroxylation of tyrosine to dopamine (DA) in intact cells, an index of endogenous TH activity, increases in parallel with the rise in TH levels. The TH mRNA level is elevated before the increase in protein levels. As determined by nuclear run‐on assays, TH gene transcription is stimulated two‐ to threefold within 30 min of addition of 1 μMDMPP to the cells; transcription returns to basal levels by 2 h. Nitrendipine (20 μM) blocks the stimulation of transcription by DMPP. Pretreatment of the cells with cycloheximide (5 μM) does not prevent the DMPP stimulation of transcription. Forskolin (10 μM) also increases TH transcription (fourfold in 15 min) by a mechanism that is not blocked by cycloheximide. These results show that nicotinic receptor stimulation increases TH mRNA synthesis, TH protein levels, and TH activity in a calcium‐dependent manner. Furthermore, the nicotinic influence on TH gene expression does not appear to require the synthesis of a protein factor for its effects. That in situ DA synthesis rates are elevated consequent to the rise in TH levels demonstrates that TH induction serves as a mechanism for enhancing the catecholamine‐synthesizing capacity of the chromaffin cell on a long‐term basis.